The present invention relates to methods of separating and partitioning differing density phases of a multiphase fluid, and more specifically, to an improved method of separating two differing density phases of a multiphase fluid and simultaneously partitioning the phases with a thixotropic material.
In U.S. Pat. No. 3,852,194 to A. R. Zine, Jr., which has been assigned to the assignee of the present application, there is described an apparatus and method for multiphase fluid collection, separation, and partitioning. The method is described in connection with collecting fresh blood and separating the blood into a serum phase and a clotted red cell phase. The method includes the steps of: providing an open-ended container with gel-like material having a specific gravity intermediate the specific gravities of two phases of a multiphase fluid to be separated; vacuum-sealing the open end of the container with a needle pierceable closure; drawing a specimen of the multiphase fluid through the closure; applying centrifugal force to the specimen and gel-like material and simultaneously forcibly moving the phases of the specimen and the gel-like material toward relative postiions within the container corresponding to their respective densities or specific gravities; terminating the application of centrifugal force after the specimen has separated into the differing density phases and a substantial portion of the gel-like material has reached a position intermediate the phases; and, at such position utilizing the gel-like material to partition the separated differing-density phases.
In one embodiment of the method of the Zine patent, the gel-like material is initially positioned adjacent the bottom end of the container such that the material is evenly distributed about the axis of the container. In particular, the gel-like material is show to be positioned adjacent a closed end of a cylindrical tubular container such that the upper surface of the gel-like material substantially lies in a plane which is normal to the axis of the tubular container. In another embodiment of the method of U.S. Pat. No. 3,852,194, the gel-like material is initially positioned on a portion of a sidewall of a tube at a distance well apart from the closed end of the tube and relatively close to the open upper end of the tube. More specifically, the material is positioned on the sidewall above the eventual interface between the separated phases.
The gel-like material utilized in the method and apparatus of the Zine patent is preferably thixotropic in its flow behavior. That is, it is substantially nonflowable at rest before centrifugation, becomes flowable during centrifugation to thereby move under the influence of centrifugation, and after centrifugation becomes substantially nonflowable again. Known types of thixotropic gel-like materials are disclosed in said Zine patent and include various mixtures of oil and finely divided siliceous dioxide. Such gels have the desirable characteristics of remaining relatively stable durig handling and storage before use and of quickly reverting to their initial nonflowable state upon termination of centrifugation.
Although the method of separating and partitioning fluid which is disclosed in the Zine patent has been satisfactory for general purposes of multiphase fluid separation, it has been found that problems may be encountered relative to the initial placement of the gel-like material specified and shown in the patent. When a thixotropic gel-like material is initially positioned at the bottom end of the tube such that the material is evenly distributed about the axis of the tube, it has been noticed that occasionally the gel-like material will remain at its initial position during normal laboratory centrifugation speeds. Even though the gel-like material has a specific gravity less than the heavier phase of the differing density fluid, nevertheless application of a normal amount of centrifugal force and even an incremental increased amount of centrifugal force sometimes did not initiate flow of the gel-like material from its initial bottom-center position.
Also, in connection with the separation of blood into a heavy phase including clotted red cells and a light phase consisting of serum, it has been found that flow of the gel-like material through central portions of the blood results occasionally in excessively high lactic dehydrogenase (LDH) content of the separated serum. As is clearly shown in the drawings of the Zine patent, initial positioning of the gel-like material at the bottom of the tube symmetrically about the axis of the tube results in flow of the gel-like material through central portions of the multiphase fluid. Although the hydrodynamics of the flow of multiphase fluids and gel-like material during centrifugation are not fully understood by the applicants, hereof, it has been determined that the flow of gel-like material from a position below a body of red cells within the tube through central portions of such body of red cells results in excessive rupturing of the red cells and the consequent release of LDH from the red cells.
Another problem associated with the positioning of the thixotropic partitioning material symmetrically at the bottom of the container relates to the quality of the seal produced by the thixotropic material between the separated phases. The movement of some thixotropic materials from such a position has been found to be sometimes so sluggish that, unless the duration or speed of centrifugation is increased, the thixotropic material may not form an adequate seal between the separated phases. By an adequate seal, it is meant that the seal must be able to remain intact while the lighter, uppermost phase is being poured or decanted from the container. It will be appreciated that an increase in the speed of centrifugation or the duration of centrifugation over that customarily used in the separation of blood is not desirable because such increases are likely to cause excessive hemolysis or rupturing of the cellular constituents of blood.
Regarding the positioning of thixotropic fluid at the sidewall of the container above the eventual interface between the separated light and heavy phases, we have found that such positioning is difficult to maintain, because thixotropic partitioning material positioned on the sidewall tends to slump and spread out over the sidewalls during handling and shipping prior to centrifugation. Thus, such elevated positioning on the sidewall produces an unprofessional appearance.
The method of the present invention responds to each of the previously described problems in a manner so as to completely eliminate any further concern regarding such problems.